Fetal hemoglobin (Hb F) is present at birth, gradually decreases in the first months of life, and normally makes up less than 2% of total hemoglobin in adults. The normal adult hemoglobin molecule (Hb A; .alpha..sub.2 .beta..sub.2) consists of two pairs of polypeptide chains designated and .beta.. In fetal hemoglobin (.alpha..sub.2 .gamma..sub.2), .gamma. chains are substituted for the .beta. chains. The types of hemoglobin chains and the chemical structure of individual polypeptides in the chains are controlled genetically. Genetic defects may result in hemoglobin molecules with abnormal physical or chemical properties, some of which may result in anemia. Such anemias, termed hemoglobinopathies, are severe in homozygotes and relatively mild in heterozygous carriers. The abnormal hemoglobins are distinguished by their electrophoretic mobility and have been designated by letters. The first to be discovered was sickle cell hemoglobin (Hb S). The important hemoglobinopathies in the Untied States are those due to Hb S, Hb C, the thalassemias, and combinations thereof.
In sickle cell anemia (Hb S Disease), valine is substituted for glutamic acid in the sixth amino acid of the .beta. chain. As a result, deoxy-Hb S is much less soluble than the normal deoxy-Hb A, and the patient's red blood cells (erythrocytes) become sickle-shaped at sites of low oxygen concentration. The distorted erythrocytes are unable to pass through small arterioles and capillaries, and plugs of such sickled erythrccytes lead to thrombosis and infarction. This leads to crises of pain which last for several days. Sickled erythrocytes are also more fragile than normal erythrocytes and less able to withstand the mechanical trauma caused by circulation in blood. Few homozygous patients live beyond age 40. Therapy has traditionally been symptomatic, e.g., transfusions are given for symptomatic anemia.
Hemoglobin S-C Disease is a moderately severe anemia due to an inherited abnormality of hemoglobin formation in which half the hemoglobin is the S type and half is of type C. There is no specific treatment.
The thalassemias are a group of chronic, familial, hemolytic anemias characterized by defective hemoglobin synthesis and ineffective erythropoiesis. .beta.-Thalassemia results from a decreased synthesis of .beta. polypeptide chains. Homozygotes (.beta.-thalassemia major) typically have severe anemia (Cooley's anemia) from infancy, and many such patients do not survive to puberty. Treatment is typically by chronic transfusion to suppress the abnormal hematopoiesis, coupled with splenectomy to alleviate the hypertrophic spleen growth (splenomegaly) that is often consequent to such hemoglobinopathies.
Clinical and experimental evidence suggests that stimulation of fetal hemoglobin will benefit patients with sickle cell anemia. Fetal hemoglobin in the adult appears to be restricted to a subpopulation of cells, the F-cells, which originate from the same progenitors as the normal adult erythrocytes that do not contain fetal hemoglobin. The numbers of F-cells are elevated in several hemopoietic disorders. Formation of F-cells has been explained by various hypotheses, including premature commitment of erythroid progenitors. Stamatoyannopoulos, G., and Th. Papayannopoulou, in Cellular and Molecular Regulation of Hemoglobin Switching, G. Stamatoyannopoulos and A.W. Nienhuis, Eds., Grune & Stratton, New York, 1979, pp. 323-350; Stamatoyannopoulos, G., et al., Ann. NY Acad. Sci. 1985, 445: 188-197. According to the latter hypothesis, erythroid progenitor cells have the ability to form F-cells but fail to do so when the kinetics of erythroid maturation are normal; F-cells are formed when progenitor cells are forced to become erythroblasts prematurely, as in sudden bone marrow expansion or when there is a sudden acceleration of erythroid cell maturation kinetics.
Recently, several attempts have been made to stimulate fetal hemoglobin in animals and in patients with sickle cell disease. Stimulation of F-cell formation has been achieved with 5-azacytidine, a drug that induces .gamma.-gene demethylation. DeSimone, J., et al., Proc. Natl. Acad. Sci. (USA) 1982, 79:4428-443; Torrealba de Ron, A., et al., Blood 1984, 63:201-210; Ley, T.J., et al., N. Engl. J. Med. 1982, 307:1469-1475; Ley, T.J., et al., Blood 1983, 62: 370-380; Nienhuis, A.W., et al., Ann. NY Acad. Sci. 1985, 445:198-211; Charache, S., et al., Proc. Natl. Acad. Sci. (USA) 1983, 80:4842-4846; Humphries, R.K., et al., J. Clin. Invest. 1985, 75:547-557; Dover, G.J., et al., Blood 1985, 66:527-532.Other cell cycle-specific drugs, such as hydroxyurea and cytarabine (Ara-C), have been found to stimulate F-cell formation in primates and human patients. Papayannopoulou, Th., et al., Science 1984, 224:617-619; Letvin, N.L., et al., New. Engl. J. Med. 1984, 310:869-873; Platt, O.S., et al., J. Clin. Invest. 1984, 74:652-656; Veith, R., et al., N. Engl. J. Med. 1986, 313:1571-1575; Dover, G.J., et al., Blood 1986, 67:735-738; Charache, S., et al., Blood 1987, 69:109-116. An M-stage compound, vinblastine, stimulates F-cell formation in baboons. Veith, R., et al., Br. J. Haemat. 1980, 44:535-546. The induction of fetal hemoglobin by cell cycle-specific drugs has been attributed to various mechanisms. One hypothesis is that the induced F-cell formation is due to the rapid erythroid regeneration kinetics triggered by the drug treatment.
Administration of erythropoietin in animals is associated with expansion of the erythroid progenitor and early precursor pools, with consequent stimulation of reticulocyte (young erythrocyte) production. Spivak, J.L., Intl. J. Cell Cloning 1986, 4:139-166; Papayannopoulou, Th., and C.A. Finch, J. Clin. Invest. 1972, 51:1179-1185. These effects are mainly due to the action of erythropoietin on late erythroid progenitors. Previous studies have shown that erythropoietin increases the number of fetal hemoglobin-positive colonies in human bone marrow cultures, induces fetal hemoglobin in cultures of monkey or baboon erythroid progenitors, and induces Hb C in sheep bone marrow cultures. Papayannopoulou, Th., et al., Proc. Natl. Acad. Sci. (USA) 1977, 74:2923-2927; Macklis, R.M., et al., J. Clin. Invest. 1982, 70:752; Torrealba de Ron, A., et al., Exp. Hematol. 1985, 13:919-925; Adamson, J.W., et al., Science 1973, 80:310-312; Nienhuis, A.W., et al., in Cellular and Molecular Regulation of Hemoglobin Switching, G. Stamatoyannopoulos and A.W. Nienhuis, Eds., 1978, Grune & Stratton, New York, 397-420. These in vitro results most likely reflect effects of erythropoietin on the kinetics of in vitro erythroid cell differentiation. Direct induction of fetal hemoglobin synthesis by erythropoietin is not supported by previous results. Papayannopoulou, Th., et al., Proc. Natl. Acad. Sci. (USA) 1979, 76:6420-6424.
Recently, recombinant erythropoietin has been introduced in the treatment of humans. Eschbach, J.W., et al., N. Eng. J. Med. 1987, 316:73-78; Winerals, C.G., et al., Lancet 1986, ii:1175-1178.